Modulation of physio-biochemical and photosynthesis parameters by overexpressing SbPIP2 gene improved abiotic stress tolerance of transgenic tobacco.

The present study aims to explore the potential of a plasma-membrane localized PIP2-type aquaporin protein sourced from the halophyte Salicornia brachiata to alleviate salinity and water deficit stress tolerance in a model plant through transgenic intervention. Transgenic plants overexpressing SbPIP2 gene showed improved physio-biochemical parameters like increased osmolytes (proline, total sugar, and amino acids), antioxidants (polyphenols), pigments and membrane stability under salinity and drought stresses compared to control plants [wild type (WT) and vector control (VC) plants]. Multivariate statistical analysis showed that, under water and salinity stresses, osmolytes, antioxidants and pigments were correlated with SbPIP2-overexpressing (SbPIP2-OE) plants treated with salinity and water deficit stress, suggesting their involvement in stress tolerance. As aquaporins are also involved in CO2 transport, SbPIP2-OE plants showed enhanced photosynthesis performance than wild type upon salinity and drought stresses. Photosynthetic gas exchange (net CO2 assimilation rate, PSII efficiency, ETR, and non-photochemical quenching) were significantly higher in SbPIP2-OE plants compared to control plants (wild type and vector control plants) under both unstressed and stressed conditions. The higher quantum yield for reduction of end electron acceptors at the PSI acceptor side [Φ( R0 )] in SbPIP2-OE plants compared to control plants under abiotic stresses indicates a continued PSI functioning, leading to retained electron transport rate, higher carbon assimilation, and less ROS-mediated injuries. In conclusion, the SbPIP2 gene functionally validated in the present study could be a potential candidate for engineering abiotic stress resilience in important crops.

Discontinuous L-binding motifs in the transactivation domain of the vesicular stomatitis virus P protein are required for terminal de novo transcription initiation by the L protein.

The phospho- (P) protein, the co-factor of the RNA polymerase large (L) protein, of vesicular stomatitis virus (VSV, a prototype of nonsegmented negative-strand RNA viruses) plays pivotal roles in transcription and replication. However, the precise mechanism underlying the transcriptional transactivation by the P protein has remained elusive. Here, using an in vitro transcription system and a series of deletion mutants of the P protein, we mapped a region encompassing residues 51-104 as a transactivation domain (TAD) that is critical for terminal de novo initiation, the initial step of synthesis of the leader RNA and anti-genome/genome, with the L protein. Site-directed mutagenesis revealed that conserved amino acid residues in three discontinuous L-binding sites within the TAD are essential for the transactivation activity of the P protein or important for maintaining its full activity. Importantly, relative inhibitory effects of TAD point mutations on synthesis of the full-length leader RNA and mRNAs from the 3'-terminal leader region and internal genes, respectively, of the genome were similar to those on terminal de novo initiation. Furthermore, any of the examined TAD mutations did not alter the gradient pattern of mRNAs synthesized from internal genes, nor did they induce the production of readthrough transcripts. These results suggest that these TAD mutations impact mainly terminal de novo initiation but rarely other steps (e.g., elongation, termination, internal initiation) of single-entry stop-start transcription. Consistently, the mutations of the essential or important amino acid residues within the P TAD were lethal or deleterious to VSV replication in host cells. IMPORTANCE RNA-dependent RNA polymerase L proteins of nonsegmented negative-strand RNA viruses belonging to the Mononegavirales order require their cognate co-factor P proteins or their counterparts for genome transcription and replication. However, exact roles of these co-factor proteins in modulating functions of L proteins during transcription and replication remain unknown. In this study, we revealed that three discrete L-binding motifs within a transactivation domain of the P protein of vesicular stomatitis virus, a prototypic nonsegmented negative-strand RNA virus, are required for terminal de novo initiation mediated by the L protein, which is the first step of synthesis of the leader RNA as well as genome/anti-genome.

Nutrient Composition, Physicobiochemical Analyses, Oxidative Stability and Antinutritional Assessment of Abundant Tropical Seaweeds from the Arabian Sea.

Foods enriched with nutritional compounds and biological activities, especially antioxidants, are considered healthier for human and/or animal consumption. Seaweeds are rich sources of biologically active metabolites and are used as functional foods. In this study, proximate compositions, physicobiochemical characteristics and oil oxidative stability were analyzed for 15 abundant tropical seaweeds (four green-Acrosiphonia orientalis, Caulerpa scalpelliformis, Ulva fasciata, Ulva lactuca; six brown-Iyengaria stellata, Lobophora variegate, Padina boergesenii, Sargassum linearifolium, Spatoglossum asperum, Stoechospermum marginatum; and five red-Amphiroa anceps, Grateloupia indica, Halymenia porphyriformis, Scinaia carnosa, Solieria chordalis). All seaweeds were analyzed for the proximate composition, including moisture content, ash content, total sugar content, total proteins, total lipids, crude fiber, carotenoid content, total chlorophyll content, proline, iodine content, nitrogen-free extract, total phenolic content and total flavonoid content. Green seaweeds showed higher nutritional proximate composition, followed by brown and red seaweeds. Among the different seaweeds, Ulva, Caulerpa, Sargassum, Spatoglossum and Amphiroa showed high nutritional proximate composition compared to other seaweeds. High cation scavenging, free radical scavenging and total reducing activities were observed for Acrosiphonia, Caulerpa, Ulva, Sargassum, Spatoglossum and Iyengaria. It was also observed that 15 tropical seaweeds contained negligible amounts of antinutritional compounds, including tannic acid, phytic acid, saponins, alkaloids and terpenoids. Nutritionally, green and brown seaweeds provided higher sources of energy (150-300 calories per 100 g) compared to red seaweeds (80-165 calories per 100 g). Additionally, this study also confirmed that tropical seaweeds improved the oxidative stability of food oils and, therefore, might be recommended as natural antioxidant additives. The overall results confirm that tropical seaweeds are potential sources of nutrition and antioxidants and may be explored as functional food, dietary supplementation or animal feed. Additionally, they may also be explored as food supplements for fortifying food products, as food toppings or for garnishing and seasoning foods. However, a human or animal toxicity analysis is required before any conclusive recommendation for daily food or feed intake can be made.

MicroRNA Biomarkers for Patients With Muscle-Invasive Bladder Cancer Undergoing Selective Bladder-Sparing Trimodality Treatment.

PURPOSE: Trimodality therapy with maximal transurethral resection of bladder tumor and definitive chemoradiation reserving cystectomy for salvage of local recurrence is an accepted treatment alternative to upfront cystectomy for selected patients with muscle-invasive bladder cancer. There is a need for molecular biomarkers to predict which patients will respond to bladder preservation therapy. METHODS AND MATERIALS: We sought to identify biomarkers with the ability to predict response to chemoradiation and survival after selective bladder preservation therapy in a cohort of 40 patients using a microRNA profiling approach. In vitro experiments were performed using transitional cell carcinoma lines CRL1749, HTB5, and HTB4. RESULTS: We identified a panel of microRNAs associated with overall survival in our bladder preservation cohort and in the TCGA cohort. We also identified several microRNAs, including miR-23a and miR-27a, microRNAs of the miR-23a cluster, to be suggestively associated with complete response to chemoradiation therapy. The microRNAs were significantly associated with overall survival in The Cancer Genome Atlas cohort. In vitro studies suggest that the functional roles of miR-23a and miR-27a involve targeting the SFRP1 protein, a negative regulator of the Wnt signaling pathway. The upregulation of ß-catenin in the Wnt signaling pathway mediated proliferation, migration, invasion, and sensitivity to radiation and cisplatin treatment in bladder cancer cells. CONCLUSIONS: Our results indicate that miR-23a and miR-27a act as oncomirs, and once independently validated, they may help appropriately triage selected bladder cancer patients to individualize treatment.

A dual-functional priming-capping loop of rhabdoviral RNA polymerases directs terminal de novo initiation and capping intermediate formation.

The L proteins of rhabdoviruses, such as vesicular stomatitis virus (VSV) and rabies virus (RABV), possess an unconventional mRNA capping enzyme (GDP polyribonucleotidyltransferase, PRNTase) domain with a loop structure protruding into an active site cavity of the RNA-dependent RNA polymerase (RdRp) domain. Here, using complementary VSV and RABV systems, we show that the loop governs RNA synthesis and capping during the dynamic stop-start transcription cycle. A conserved tryptophan residue in the loop was identified as critical for terminal de novo initiation from the genomic promoter to synthesize the leader RNA and virus replication in host cells, but not for internal de novo initiation or elongation from the gene-start sequence for mRNA synthesis or pre-mRNA capping. The co-factor P protein was found to be essential for both terminal and internal initiation. A conserved TxΨ motif adjacent the tryptophan residue in the loop was required for pre-mRNA capping in the step of the covalent enzyme-pRNA intermediate formation, but not for either terminal or internal transcription initiation. These results provide insights into the regulation of stop-start transcription by the interplay between the RdRp active site and the dual-functional priming-capping loop of the PRNTase domain in non-segmented negative strand RNA viruses.